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Polymersome formulations improve immunogenicity of RBD in COVID-19 vaccine
Rachel P. Wallace1, Lisa R. Volpatti*1, Shijie Cao*1, Michal M. Raczy*1, Ruyi Wang*1, L. Taylor Gray1, Aaron T. Alpar1, Priscilla S. Briquez1, Nikolaos Mitrousis1, Tiffany M. Marchell2, Maria Stella Sasso1, Aslan Mansurov1, Erica Budina1, Andrew C. Tremain2, Melody A. Swartz1, Jeffrey A. Hubbell1
1Pritzker School of Molecular Engineering, University of Chicago; 2Committee on Immunology, University of Chicago
*These authors contributed equally to this work
Since December 2019, the disease COVID-19 spread by the SARS-CoV2 virus has become a global pandemic, spurring efforts to produce an effective vaccine1. A promising antigen for vaccination is the spike protein (S), which enables SARS-CoV2 binding to ACE-2 and its infectivity. Within this protein, the receptor-binding domain (RBD), a 197 amino acid fragment of S, has been identified to directly engage ACE-22. Previously, PEG-PPS polymersomes have been used in vaccines to effectively target antigen to antigen-presenting cells in the lymph node and increase cross-presentation and CD8+ T cell activation3. In order to develop an effective vaccine against COVID-19 using RBD as an antigen, we used PEG-PPS polymersomes to investigate two different antigen formulations, encapsulated RBD (RBDencap), and surface-conjugated RBD (RBDsurf) in combination with PEG-PPS polymersome encapsulated monophosphoryl lipid A (MPLA) as an adjuvant.
We treated mice with our polymersome vaccines and boosted three weeks later. Weekly bleeding was done to measure the anti-RBD antibody response via ELISA. One week after the boost mice were sacrificed, splenocytes and lymphocytes were isolated and analyzed via flow cytometry for the presence of CD3+CD4+PD1+CXCR5+ Tfh cells and the phenotype of RBD-specific B cells identified using fluorescent RBD probes. Splenocytes and lymphocytes were restimulated using RBD and analyzed for production of proinflammatory cytokines. We used peptide arrays to determine the epitope specificity of the generated antibodies and measured naturalization capability of the antibodies against the SARS-CoV2 virus.
Both RBDencap and RBDsurf were able to generate high levels of RBD-specific IgG antibodies, Tfh cell responses, and antigen-specific CD8+ T cell responses upon restimulation compared to unformulated RBD. The RBDencap generated predominantly RBD-specific B220+CD138+ plasmablasts and mounted a very quick antibody response to a broad array of epitopes. These antibodies were not able to neutralize the SARS-CoV2 virus. In comparison, the RBDsurf generated an equally robust antibody response delayed by approximately a week and the RBD-specific B cells were predominantly B220+CD38-CL7+ germinal center B cells. The antibodies generated were specific to a more selective set of epitopes, likely due to increased somatic hypermutation, and were able to effectively neutralize SAR-CoV2. In summary, we demonstrated that while antigen formulation via PEG-PPS polymersomes can increase RBD-specific CD8+ T cell responses, only surface-conjugated RBD polymersomes can effectively generate a neutralizing antibody response against SARS-CoV2.
1.Huang, C. et al. Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet 395, 497–506 (2020).
2.Lan J. et al. Structure of the SARS-CoV-2 spike receptor-binding domain bound to the ACE2 receptor. Nature 518, 215–220 (2020).
3.Scott, E.A. et al. Dendritic cell activation and T cell priming with adjuvant- and antigen-loaded oxidation-sensitive polymersomes. Biomaterials 33, 6211-6219 (2012).